THE EFFECT OF ADMINISTRATION OF AN EQUAL DOSE OF DIFFERENT CLASSES OF PHYTOCHEMICALS ON HEME OXYGENASE-1 GENE AND PROTEIN EXPRESSION IN MICE LIVER
Objective: Heme oxygenase-1 (HO-1) is enzyme that possesses antioxidant, anti-inflammatory, and cytoprotective functions. Induction of HO-1 occurs as an adaptive and beneficial response to various injurious stimuli such as oxidative stress. This study is aimed at monitoring the effects of administration of equal doses (50 mg/kg) of sulforaphane (SFN), curcumin, quercetin, indole-3-carbinol, and butylated hydroxyanisole (BHA) for 14 days on the levels of liver HO-1 gene and protein expression in mice.
Method: A total of 48 adult male ICR white mice (25–30 g) were divided into eight groups: Normal control group (n=6), SFN-treated group (n=6), quercetin-treated group (n=6), curcumin-treated group (n=6), BHA-treated group (n=6), indole-3-carbinol treated group (n=6), vehicle 1 control group (n=6), and vehicle 2 control group (n=6). All chemicals were administered intraperitoneally at a dose of 50 mg/kg for 14 days. Vehicle 1 (dimethyl sulfoxide, TweenTM 20, and normal saline at a ratio of 0.05:0.1:0.85) was used to dissolve SFN, quercetin, and curcumin. Vehicle 2 (corn oil) was used to dissolve indole-3-carbinol and BHA. At day 15, the animals were sacrificed and their livers were isolated. From the liver, total RNA was extracted, reverse transcribed and subjected to quantitative real‐time polymerase chain reaction to detect HO-1 gene expression. Agarose gel electrophoresis was also performed to verify the specificity of the amplification. HO-1 protein expression was determined by Western blotting.
Results: HO-1 gene expression showed significant increase of 4.6±0.3, 3.6±0.2, 3.6±0.4, 3.3±0.3, and 3.0±0.4-fold and HO-1 protein expression showed significant increase of 2.3±0.2, 2.2±0.2, 2.2±0.1, 1.8±0.1, and 1.7±0.2-fold following treatment with 50 mg/kg of SFN, indole-3-carbinol, BHA, curcumin, and quercetin, respectively, compared to controls (p<0.05).
Conclusion: At a dose of 50 mg/kg, SFN administration for 14 days resulted in the highest induction of HO-1 gene and protein expression level in mice liver, and quercetin the lowest.
2. Abuajah CI, Ogbonna AC, Osuji CM. Functional components and medicinal properties of food: A review. J Food Sci Technol 2015;52:2522-9.
3. Keum YS. Regulation of nrf2-mediated phase II detoxification and anti-oxidant genes. Biomol Ther (Seoul) 2012;20:144-51.
4. Kitteringham NR, Abdullah A, Walsh J, Randle L, Jenkins RE, Sison R, et al. Proteomic analysis of nrf2 deficient transgenic mice reveals cellular defence and lipid metabolism as primary nrf2-dependent pathways in the liver. J Proteomics 2010;73:1612-31.
5. Abdullah A, Kitteringham NR, Jenkins RE, Goldring C, Higgins L, Yamamoto M, et al. Analysis of the role of nrf2 in the expression of liver proteins in mice using two-dimensional gel-based proteomics. Pharmacol Rep 2012;64:680-97.
6. He CH, Gong P, Hu B, Stewart D, Choi ME, Choi AM, et al. Identification of activating transcription factor 4 (ATF4) as an nrf2- interacting protein. Implication for heme oxygenase-1 gene regulation. J Biol Chem 2001;276:20858-65.
7. Keum YS, Han YH, Liew C, Kim JH, Xu C, Yuan X, et al. Induction of heme oxygenase-1 (HO-1) and NAD[P]H: Quinone oxidoreductase 1 (NQO1) by a phenolic antioxidant, butylated hydroxyanisole (BHA) and its metabolite, tert-butylhydroquinone (tBHQ) in primary-cultured human and rat hepatocytes. Pharm Res 2006;23:2586-94.
8. Nussler AK, Hao L, Knobeloch D, Yao P, Nussler NC, Wang Z, et al. Protective role of HO-1 for alcohol-dependent liver damage. Dig Dis 2010;28:792-8.
9. Domitrovi? R, Jakovac H, Vasiljev Marchesi V, Vladimir-Kneževi? S, Cvijanovi? O, Tadi? Z, et al. Differential hepatoprotective mechanisms of rutin and quercetin in CCl(4)-intoxicated BALB/cN mice. Acta Pharmacol Sin 2012;33:1260-70.
10. Noh JR, Kim YH, Hwang JH, Choi DH, Kim KS, Oh WK, et al. Sulforaphane protects against acetaminophen-induced hepatotoxicity. Food Chem Toxicol 2015;80:193-200.
11. Ji LL, Sheng YC, Zheng ZY, Shi L, Wang ZT. The involvement of p62- keap1-nrf2 antioxidative signaling pathway and JNK in the protection of natural flavonoid quercetin against hepatotoxicity. Free Radic Biol Med 2015;85:12-23.
12. Hajra S, Patra AR, Basu A, Bhattacharya S. Prevention of doxorubicin (DOX)-induced genotoxicity and cardiotoxicity: Effect of plant derived small molecule indole-3-carbinol (I3C) on oxidative stress and inflammation. Biomed Pharmacother 2018;101:228-43.
13. Liu S, Tian L, Chai G, Wen B, Wang B. Targeting heme oxygenase-1 by quercetin ameliorates alcohol-induced acute liver injury via inhibiting NLRP3 inflammasome activation. Food Funct 2018;9:4184-93.
14. Luo L, Chen Y, Wu D, Shou J, Wang S, Ye J, et al. Butylated hydroxyanisole induces distinct expression patterns of nrf2 and detoxification enzymes in the liver and small intestine of C57BL/6 mice. Toxicol Appl Pharmacol 2015;288:339-48.
15. Liu XM, Azam MA, Peyton KJ, Ensenat D, Keswani AN, Wang H, et al. Butylated hydroxyanisole stimulates heme oxygenase-1 gene expression and inhibits neointima formation in rat arteries. Cardiovasc Res 2007;74:169-79.
16. Sharma A, Sharma MK, Kumar M. Protective effect of Mentha piperita against arsenic-induced toxicity in liver of swiss albino mice. Basic Clin Pharmacol Toxicol 2007;100:249-57.
17. Sanghamitra S, Hazra J, Upadhyay SN, Singh RK, Amal RC. Arsenic induced toxicity on testicular tissue of mice. Indian J Physiol Pharmacol 2008;52:84-90.
18. Sharma A, Sharma MK, Kumar M. Modulatory role of emblica officinalis fruit extract against arsenic induced oxidative stress in Swiss albino mice. Chem Biol Interact 2009;180:20-30.
19. Roehr B. Why sex matters in mouse models. Sci 2007;21:49.
20. Molina MF, Sanchez-Reus I, Iglesias I, Benedi J. Quercetin, a flavonoid antioxidant, prevents and protects against ethanol-induced oxidative stress in mouse liver. Biol Pharm Bull 2003;26:1398-402.
21. Innamorato NG, Rojo AI, García-Yagüe AJ, Yamamoto M, de Ceballos ML, Cuadrado A, et al. The transcription factor nrf2 is a therapeutic target against brain inflammation. J Immunol 2008;181:680-9.
22. Girish C, Koner BC, Jayanthi S, Ramachandra Rao K, Rajesh B, Pradhan SC, et al. Hepatoprotective activity of picroliv, curcumin and ellagic acid compared to silymarin on paracetamol induced liver toxicity in mice. Fundam Clin Pharmacol 2009;23:735-45.
23. Priya DK, Gayathri R, Gunassekaran GR, Sakthisekaran D. Protective role of sulforaphane against oxidative stress mediated mitochondrial dysfunction induced by benzo(a)pyrene in female Swiss albino mice. Pulm Pharmacol Ther 2011;24:110-7.
24. Oh CJ, Kim JY, Min AK, Park KG, Harris RA, Kim HJ, et al. Sulforaphane attenuates hepatic fibrosis via NF-E2-related factor 2-mediated inhibition of transforming growth factor-?/Smad signaling. Free Radic Biol Med 2012;52:671-82.
25. Heeba GH, Mahmoud ME, El Hanafy AA. Anti-inflammatory potential of curcumin and quercetin in rats: Role of oxidative stress, heme oxygenase-1 and TNF-?. Toxicol Ind Health 2014;30:551-60.
26. Livak KJ, Schmittgen TD. Analysis of relative gene expression data using real-time quantitative PCR and the 2(-delta delta C(T)) method. Methods 2001;25:402-8.
27. Chang M, Xue J, Sharma V, Habtezion A. Protective role of hemeoxygenase-1 in gastrointestinal diseases. Cell Mol Life Sci 2015;72:1161-73.
28. Kandavelu S, Somasundram PC, John B, Rajendran R. Pro-inflammatory cytokines elicit inflammatory response in blood leukocytes of post dialytic chronic renal patients through heme oxygenase-1 activation. Int J Pharm Pharm Sci 2014;6:111-5.
29. Choi JH, Kim DW, Yun N, Choi JS, Islam MN, Kim YS, et al. Protective effects of hyperoside against carbon tetrachloride-induced liver damage in mice. J Nat Prod 2011;74:1055-60.
30. Eipel C, Hirschmann M, Abshagen K, Menger MD, Vollmar B. Local interaction of apoptotic hepatocytes and kupffer cells in a rat model of systemic endotoxemia. Hepatol Res 2007;37:863-71.
31. Brockmann JG, August C, Wolters HH, Hömme R, Palmes D, Baba H, et al. Sequence of reperfusion influences ischemia/reperfusion injury and primary graft function following porcine liver transplantation. Liver Transpl 2005;11:1214-22.
32. Martin D, Rojo AI, Salinas M, Diaz R, Gallardo G, Alam J, et al. Regulation of heme oxygenase-1 expression through the phosphatidylinositol 3-kinase/Akt pathway and the nrf2 transcription factor in response to the antioxidant phytochemical carnosol. J Biol Chem 2004;279:8919-29.
33. Cho HY, Jedlicka AE, Reddy SP, Kensler TW, Yamamoto M, Zhang LY, et al. Role of NRF2 in protection against hyperoxic lung injury in mice. Am J Respir Cell Mol Biol 2002;26:175-82.
34. Saka S, Singh AN, Sharma N. Potential anti-cancer superfoods: A minireview. Int J Curr Pharm Res 2016;8:19-21.
35. Islam MT, De Alencar MV, Paz MF, Matos LA, Sousa JM, Melo- Cavalcante AA. Coffee: A health fuel-blot popular drinking. Int J Pharm Pharm Sci 2016;8:1-7.
36. Yao P, Nussler A, Liu L, Hao L, Song F, Schirmeier A, et al. Quercetin protects human hepatocytes from ethanol-derived oxidative stress by inducing heme oxygenase-1 via the MAPK/Nrf2 pathways. J Hepatol 2007;47:253-61.
37. Bao W, Li K, Rong S, Yao P, Hao L, Ying C, et al. Curcumin alleviates ethanol-induced hepatocytes oxidative damage involving heme oxygenase-1 induction. J Ethnopharmacol 2010;128:549-53.
38. Liu S, Hou W, Yao P, Li N, Zhang B, Hao L, et al. Heme oxygenase-1 mediates the protective role of quercetin against ethanol-induced rat hepatocytes oxidative damage. Toxicol In Vitro 2012;26:74-80.
39. Sass G, Barikbin R, Tiegs G. The multiple functions of heme oxygenase-1 in the liver. Z Gastroenterol 2012;50:34-40.
40. Liu B, Qian JM. Cytoprotective role of heme oxygenase-1 in liver ischemia reperfusion injury. Int J Clin Exp Med 2015;8:19867-73.
41. Khan YY, Suvarna V. Liposomes containg phytochemicals for cancer treatment - An update. Int J Curr Pharm Res 2017;9:20-4.
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